A THERMODYNAMIC ANALYSIS OF THE SEQUENCE-SPECIFIC BINDING OF RNA BY BACTERIOPHAGE-MS2 COAT PROTEIN

Most mutations in the sequence of the RNA hairpin that specifically hi nds MS2 coat protein either reduce the binding affinity or hare no eff ect. However, one RNA mutation, a uracil to cytosine change ire the lo op, has the unusual property of increasing the binding affinity to the protein by nearly 100-fold. Guided by the structure of the protein-RN A complex, we used a series of protein mutations and RNA modifications to evaluate the thermodynamic basis for the improved affinity: The ti ght binding of the cytosine mutation is due to (i) the amino group of the cytosine residue making an intra-RNA hydrogen bond that Increases the propensity of the free RNA to adopt the structure seen in the comp lex and (ii) the increased affinity of hydrogen bonds between the prot ein and a phosphate two bases away from the cytosine residue. The data are in good agreement with a recent comparison of the cocrystal struc tures of the two complexes, where small differences in the two structu res are seen at the thermodynamically important sites.